To further characterize the properties of retinal horizontal cell electrical synapses, we have studied the gating characteristics of gap junctions between cone-driven horizontal cells from the hybrid striped bass retina using double whole-cell voltage-clamp techniques. In a total of 105 cell pairs, the macroscopic conductance ranged from 0.4-100 nS with most cell pairs exhibiting junctional conductances between 10 and 30 nS. The junctional current-voltage relationship showed that peak or instantaneous currents (Iinst) were linear within the Vj range of +/- 100 mV and that steady-state junctional currents (Iss) exhibited rectification with increasing voltage beginning around +/- 30-40 mV Vj. The normalized junctional current-voltage relationship was well fit by a two-state Boltzmann distribution, with an effective gating charge of 1.9 charges/channel, a half-maximal voltage of approximately +/- 55 mV, and a normalized residual conductance of 0.28. The decay of junctional current followed a single exponential time course with the time constant decreasing with increasing Vj. Recovery of junctional current from voltage-dependent inactivation takes about 1 s following a pulse of 80 mV, and is about five times slower than the inactivation time course at the same Vj. Single-channel analysis showed that the unitary conductance of junctional channels is 50-70 pS. The overall open probability decreased in a voltage-dependent manner. Both the mean channel open time and the frequency of channel opening decreased, while the channel closure time increased. The ratio of closed time/total recording time significantly increased as Vj increased. Increased Vj reduced the number of events at all levels and shifted the unitary conductance to a lower level. Kinetic analysis of channel open duration showed that the distribution of channel open times was best fit by two exponentials and increased Vj significantly reduced the slower time constant. These results indicate that bass retina horizontal cells exhibit voltage-dependent inactivation of macroscopic junctional current. The inactivation occurs at the single-channel level mainly by increasing the rate of closure of voltage-sensitive channels.

Animals↗

PubMedDate not supplied

Electrical coupling of retinal horizontal cells mediated by distinct voltage-independent junctions.

Electrical coupling between H2 horizontal cell pairs isolated from the hybrid bass retina was studied using dual whole-cell, voltage-clamp technique. Voltage-dependent inactivation of junctional currents in response to steps in transjunctional voltage (Vj) over a range of +/-100 mV was characterized for 89 cell pairs. Approximately one-quarter of the pairs exhibited strongly voltage-dependent junctions (>50% reduction in junctional current at +/-100 mV), another quarter of the pairs exhibited voltage-independent junctional current (<5% reduction at +/-100 mV), and the remainder of the pairs exhibited intermediate values for voltage inactivation. We focused on further characterizing the Vj-independent junctions of horizontal cells, which have not been described previously in detail. When Lucifer Yellow dye was included in one recording pipette, pairs exhibiting Vj-independent coupling showed no (9/12), or limited (3/12), passage of dye. Vj-independent coupling was markedly less sensitive to the modulators SNP (100-300 microM, -9% reduction in coupling) and dopamine (100-300 microM, -6%) than were Vj-dependent junctions (-45% and -44%). However, simultaneous application of both SNP and dopamine significantly reduced Vj-independent coupling (-56%). Both Vj-independent and Vj-dependent junctions were blocked by DMSO (1-2%), but Vj-independent junctions were not blocked by heptanol. Single-channel junctional conductances of Vj-independent junctions range from 112-180 pS, versus 50-60 pS for Vj-dependent junctions. The results reveal that Vj-independent coupling in a subpopulation of horizontal cells from the hybrid bass retina is mediated by cellular junctions with physiological and pharmacological characteristics distinct from those previously described in fish horizontal cells.

Animals↗

PubMedDate not supplied

Strategies for assessing children's organophosphorus pesticide exposures in agricultural communities.

Children can be exposed to pesticides from multiple sources and through multiple pathways. In addition to the standard pathways of diet, drinking water and residential pesticide use, children in agricultural communities can be exposed to pesticides used in agricultural production. A research program on children and pesticides was established at the University of Washington (UW) in 1991 and has focused on two major exposure pathway issues: residential proximity to pesticide-treated farmland and transfer of pesticides from the workplace to the home (paraoccupational or take-home exposure). The UW program selected preschool children of agricultural producers and farm workers in the tree fruit region of Washington state as a population that was likely to have elevated exposures from these pathways. The organophosphorus (OP) pesticides were selected as a common class of chemicals for analysis so that issues of aggregate exposure and cumulative risk could be addressed. This paper provides an overview of key findings of our research group over the past 8 years and describes current studies in this field. Soil and housedust concentrations of OP pesticides were elevated in homes of agricultural families (household members engaged in agricultural production) when compared to non-agricultural reference homes in the same community. Dialkyl phosphate metabolites of OP pesticides measured in children's urine were also elevated for agricultural children when compared to reference children and when compared to children in the Seattle metropolitan area. Proximity to farmland was associated with increased OP pesticide concentrations in housedust and OP pesticide metabolites in urine. Current studies include a community-based intervention to reduce parental transfer of pesticides from the workplace, and a systematic investigation of the role of agricultural spray drift in children's exposure to pesticides.

Agriculture↗

PubMedDate not supplied

An ergonomic analysis of premixing and compounding processes in an animal health plant.

This study is based on an ergonomic job analysis designed to develop a hazard prevention program for the premixing and compounding processes in animal health products plants. Animal health products plants are "pharmaceutical facilities" for livestock or domestic animals. A hazardous aerosol (highly concentrated antibiotics, anthelmintics, mineral oil, and animal hormones) is generated in the premixing or compounding processes. The animal health premixing jobs are heavy-duty jobs and have high potential for chemical exposure, heat stress, and ergonomic hazards. Ergonomic job analysis was used to recognize, identify, and evaluate actuarial and potential risks of injures or irritations. Chemical hazards and eight ergonomic factors were discussed: (1) forceful exertions, (2) awkward postures, (3) localized contract stresses, (4) vibration, (5) noise, (6) temperature extremes, (7) repetitive activities, and (8) prolonged activities. The results show that (1) current practices do not violate occupational safety and health regulations or recommended guidelines, but that hazards should be identified to protect worker health and safety; (2) for chemical hazards prevention, operators wear whole-body protection equipment, which also causes heat stress and increases the noise level in the work zone; and (3) the loading weight of the bags used needs to be reduced.

Air Pollution, Indoor↗

PubMedDate not supplied

Immunomagnetic separation of adult human olfactory neural progenitors.

Olfactory neuroepithelium (ONe) has lifelong regenerative capacity owing to the presence of mitotically active progenitors. The accessibility of ONe makes it a unique source of progenitors for cell replacement strategies in the CNS. We have established lines of neurosphere forming cells (NSFCs) from adult postmortem ONe and patients undergoing nasal sinus surgery by endoscopic biopsy. These heterogeneous lines are composed primarily of an immature neuronally restricted and a small glial restricted subpopulation. More homogeneous subpopulations of the NSFCs are essential for detailed study of factors influencing their lineage restriction. Immunomagnetic bead separation using an antibody against tyrosine kinase (Trk) receptors (Trk-pan, which recognizes Trk-A, B, C) resulted in viable, enriched positive and negative subpopulations that could be analyzed immunocytochemically. The positive cells remained positive for the first week after which the number of Trk-pan expressing cells decreased. The negative subpopulation began to express Trk-pan immunoreactivity after five days in vitro. Both subpopulations reverted to the heterogeneous composition after two weeks. Furthermore, most NSFCs were positive for Trk-B, a few for Trk-A, while no reactivity was observed for Trk-C. Because NSFCs produce brain derived neurotrophic factor (BDNF) and express Trk B, the specific receptor for BDNF, it is likely that population dynamics are under a paracrine and/or autocrine regulatory mechanism. Lineage restriction analysis demonstrated that the isolated subpopulation had a restriction potential equivalent to the original heterogeneous population. These studies characterize further the NSFCs and support the future potential therapeutic use of ONe-derived progenitors for CNS injury and neurodegenerative disorders.

Adult↗

PubMedDate not supplied

Clonal analysis of adult human olfactory neurosphere forming cells.

Olfactory neuroepithelium (ONe) is unique because it contains progenitor cells capable of mitotic division that replace damaged or lost neurons throughout life. We isolated populations of ONe progenitors from adult cadavers and patients undergoing nasal sinus surgery that were heterogeneous and consisted of neuronal and glial progenitors. Progenitor lines have been obtained from these cultures that continue to divide and form nestin positive neurospheres. In the present study, we used clonal and population analyses to probe the self-renewal and multipotency of the neurosphere forming cells (NSFCs). NSFCs plated at the single cell level produced additional neurospheres; dissociation of these spheres resulted in mitotically active cells that continued to divide and produce spheres as long as they were subcultured. The mitotic activity of clonal NSFCs was assessed using bromodeoxyuridine (BrdU) incorporation. Lineage restriction of the clonal cultures was determined using a variety of antibodies that were characteristic of different levels of neuronal commitment: ss-tubulin isotype III, neural cell adhesion molecule (NCAM) and microtubule associated protein (MAP2), or glial restriction: astrocytes, glial fibrillary acidic protein (GFAP); and oligodendrocytes, galactocerebroside (GalC). Furthermore, nestin expression, a marker indicative of progenitor nature, decreased in defined medium compared to serum-containing medium. Therefore, adult human ONe-derived neural progenitors retain their capacity for self-renewal, can be clonally expanded, and offer multipotent lineage restriction. Therefore, they are a unique source of progenitors for future cell replacement strategies in the treatment of neurotrauma and neurodegenerative diseases.

Adult↗

PubMedDate not supplied

The dynamic distribution of fluoro-gold and its interrelation with neural nitric oxide synthase following intracerebroventricular injection into rat brain.

We mapped the dynamic distribution of fluoro-gold (FG) within rat brain following intracerebroventricular (icv) injection into the lateral ventricle and observed its interrelation with neural nitric oxide synthase (nNOS) using FG fluorescent microphotography combined with nNOS immunohistochemistry. We also detected the amount of icv administered FG entering the peripheral circulation using a fluorescence microplate assay. The degree of periventricular penetration of FG was significantly increased over time. At 2 min after icv injection, FG primarily labeled the choroid plexus in the lateral and third ventricles, with limited penetration into the ependyma and the subependyma of the same ventricles. Some FG/nNOS-double labeled cerebrospinal fluid-contacting neurons were observed in these ventricles as well. At 15 and 30 min, FG penetrated mainly into forebrain ventricular organs and parenchymal structures. Many FG/nNOS double labeled neurons were found at each of these sites. In addition, at 30 min intense FG labeling was found in the hypophysis, while limited periventricular penetration of FG was detected in the hindbrain circumventricular areas. In the peripheral circulation, a low concentration of FG was detected 2 min after icv injection. The concentration increased slowly, peaked at 20 min, then gradually decreased until the end of the experiment at 30 min. These findings indicate that dynamic penetration of icv administrated agents into the periventricular tissues and peripheral circulation should be considered when designing icv experiments.

Animals↗

PubMedDate not supplied

Improved cleanup and determination of dialkyl phosphates in the urine of children exposed to organophosphorus insecticides.

Analysis of dialkylphosphate urinary metabolites of organophosphorus insecticides has been used to estimate dose in nonoccupationally exposed populations, including children. Analytical methods must continue to be improved in order to accurately and reproducibly measure less than 10 ng/mL of these metabolites. Dialkyl phosphates are commonly determined as their pentafluorobenzyl bromide derivatives via gas chromatography (GC) with flame photometric detection. Presented here is an improved method for precleanup of urine using solid-phase extraction, followed by derivatization and GC analysis. The method includes the quantitative determination of the following dialkyl phosphate metabolites: dimethylphosphate, diethylphosphate, dimethylthiophosphate, diethylthiophosphate, and dimethyldithiophosphate. Additional cleanup of urine samples allows for increasing sample size and improving sensitivity while minimizing interferences and variability associated with derivatization. Sample aliquot size was 5 mL with limits of quantitation of 10 ng/mL of urine for dimethylphosphate and diethylphosphate and 2 ng/mL of urine for dimethylthiophosphate, diethylthiophosphate, and dimethyldithiophosphate. This level of method sensitivity allows for quantitative determination of trace dialkyl phosphates in approximately 75% of individuals in nonoccupationally exposed populations. This streamlined method increases sample throughput, provides a clean extract for analysis, and requires no custom glassware.

Alkylation↗

PubMedDate not supplied

Association between tumor angiogenesis and Borrmann type 4 carcinomas of the stomach.

A total of 202 gastric carcinomas, including 33 type 4 carcinomas, were studied. Intratumoral vessels were stained with an anti-CD34 monoclonal antibody before being quantitated by light microscopy (x200). The mean vessel counts for type 4 carcinomas were significantly higher than those for any other type of carcinoma, with the exception of type 1 carcinomas. Multivariate logistic regression showed that the vessel count was one of independent variables associated with type 4 carcinomas. In addition, survival analysis demonstrated that patients with type 4 carcinoma had significantly less favorable survival rates than those with other types of carcinomas.

Aged↗

PubMedDate not supplied

Immunosuppressive effect of 2'-deoxycoformycin (Pentostatin) for rat islet allotransplantation.

Adenosine deaminase (ADA) is an important enzyme for proper function of lymphocytes and congenital absence of ADA results in a form of severe combined immunodeficiency syndrome. 2'-Deoxycoformycin (Pentostatin, DCF) irreversibly inhibits ADA and therefore has been suggested as an immunosuppressive drug. The present study evaluated the immunosuppressive effect of DCF for islet allotransplantation in rats. Isolated islets (1,500 islets) from Lewis rats were transplanted into the kidney subcapsular space of streptozotocin-induced diabetic Wistar-Furth rats. DCF was administered IP either as a single injection at 1 mg/kg/wk, 1 mg/kg twice weekly, 5 mg/kg/twice weekly or 1 mg/kg/day, or as a continuous infusion at 0.8 or 1 mg/kg/day. Daily administration of DCF at 0.8 mg/kg in both methods, single daily injection or continuous infusion, resulted in a lymphopenia and a decrease in concanavalin A stimulation of splenic lymphocytes. However, DCF (in all doses) was not effective in preventing islet allograft rejection as evaluated by measuring the duration of normoglycemia following islet transplantation and by microscopic examination of the islet grafts. In fact, the duration of normoglycemia following islet transplantation was 7.5 +/- 0.9 and 9.0 +/- 1.0 days in rats receiving DCF in single daily injection or continuous infusion, respectively. This was not significantly different from control nontreated transplanted rats (8.5 +/- 0.7 days). Increasing the dose of DCF to 1 mg/kg, administered by continuous infusion, resulted in 100% mortality. For comparison, cyclosporine A (20 mg/kg, IP daily injection for 14 days) prolonged islet allograft survival to 27.3 +/- 1.5 days (p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMedDate not supplied

Ultrasonographic findings of papillary thyroid carcinoma and their relation to pathologic changes.

To elucidate the ultrasonographic findings in papillary thyroid carcinoma and their relation to pathologic changes, 47 patients on whom thyroid ultrasonography had been performed and whose thyroid tissues were pathologically proven to be papillary thyroid carcinoma were studied. The ultrasonographic findings were reviewed and correlated with the pathologic changes. Fifty-three out of 94 thyroid lobes had papillary carcinoma. The ultrasonographic features of the affected thyroid lobes were hypoechoic in 46, isoechoic in 6 and hyperechoic in 1. The ultrasonographic texture of all 53 lesions was heterogeneous. The margin was clear in 11 lesions and unclear in 42. Cystic degeneration was found in 15 lesions. Discrete particles representative of microcalcifications were found in 25 lesions. Halo signs were found in 7 lesions. Lymph node enlargement was detected in 4 cases. The sensitivity and specificity of each ultrasonic finding in predicting the respective pathologic feature were: unclear margin for tumor invasion, 84% and 31%; cystic degeneration for cysts, 42% and 79%; discrete particles for microcalcification, 50% and 52%; halo sign for total encapsulation, 42% and 88%; and lymph node enlargement for lymph node metastasis, 18% and 100%, respectively. These data suggest that ultrasonographic findings in papillary carcinoma were usually hypoechoic and heterogeneous. An unclear margin in sonography is fairly sensitive for the prediction of tumor invasion.

Adolescent↗